104 THE BACTERIOPHAGE AND ITS BEHAVIOR 



2. FIXATION OF THE BACTERIOPHAGE CORPUSCLE 



In view of the fact that it is possible to enumerate the bacteriophage 

 corpuscles present in a liquid, we can now go somewhat further into the 

 mechanism of the phenomenon of bacteriophagy. 



One of the first questions to consider bears upon the sphere of activ- 

 ity of the bacteriophage. If, in an appropriate liquid, we combine 

 bacteriophage corpuscles and susceptible bacteria, do the corpuscles 

 act at a distance or must they first come into immediate contact with 

 the bacterial cell? 



The following experiment (d'Herelle^^^) answers this question. 



The following suspensions are prepared: 



1. One hundred cubic centimeters of a suspension of the Shiga bacil- 

 lus containing 250 million bacilli per cubic centimeter. This is inocu- 

 lated with 0.25 cc. of bacteriophage suspension. 



2. One hundred cubic centimeters of a suspension of the cholera 

 vibrio, containing 250 million bacilli per cubic centimeter. This also 

 is inoculated with 0.25 cc. of the same suspension of Shiga-bacteriophage. 



3. One hundred cubic centimeters of bouillon containing only 0.25 

 cc. of the same bacteriophage. 



The material of all three flasks is incubated at 37°C. Immediately 

 after the inoculation, after 30 minutes, and again after 1 hour, 20 cc. 

 are taken from each of the three flasks and centrifuged at 4000 revolu- 

 tions per minute for 10 minutes. 



There are thus 9 tubes which have been centrifuged. From the 

 supernatant fluid of each of these, 0.02 cc. is taken and introduced into 

 other tubes containing suspensions of the Shiga bacillus, and counts 

 of the corpuscles are made by plating 0.02 cc, of each of these 9 tubes 

 on six plates of medium. In this way an average of the counts can be 

 obtained, and the results of the counts indicate the number of corpuscles 

 remaining in the medium, since those which have penetrated the bac- 

 terial cells before the centrifugation have been thrown down with the 

 cells during this procedure and, as a result are to be found in the sedi- 

 ment. 



The results of the counts are as follows: 



Tube 1. Shiga suspension plus bacteriophage. 



a. Counts of the material made immediately after the preparation 

 are 214, 193, 187, 221, 229, and 183 plaques. The average is 204, 

 representing 5,000,000 corpuscles per cubic centimeter in the original 

 suspension immediately after inoculation. 



